Choline ascorbate, methods for producing same, and compositions thereof



United States Patent-O CHOLINE ASCORBATE', METHODS FORPRO'DUC INGQSAME, AND COMPOSITIONS THEREOFN Walter'H. Hoffman; s rin fieimt/ie.

-. No"D'rawing.e Application. November 17,1954 Serial-No. 4695601 12 Claims. (Cl."167-81) This? inventionrelates. to a novel derivative of ascorbic acid, and, more particularly, to a choline derivative .of levo-ascorbic l acid lhereinafter designated..-as-choline ascorbate. The invention furthenrelates' to methods fo'r preparing r choline. ascorbate," and 'to" stableTtherap'eutic or nutrient compositions containingcholine asc'orbate Ascorbic acid, better known-*asuvitam-ineC, is :oneof the essential dietary factors and is frequently. applied in the fieldsof nutrition and medicine.' However, it suffers the disadvantage of being easily oxidized and is relatively unstable on storage. Moreover, ascorbic-acid has. been fOUl'ldrttO .be incompatible-with certain other vitamins and therapeutic agents with which it would otherwise provide a 'desirablecombination for supplying several dietary or therapeutic needs. It has beensubstantially impossible to prepare a suitable parenteral solution of ascorbic acid either alone or in combination with other vitamins: For example-"parenteral preparations containing'y'itamin" B 12 and ascorbic acid" were found tobe unstable"and-"incompatiblc, resulting in dysfunction of the former" and rapid loss ofiactivit'y of both con stituents;

Cholineis' also an essential dietary 'co' rnponentaan'd is often'clas'sified'as 'a'member'of theB -co'rnplex group of vitamins. Its chief physiological function is predicated on the fact that it is theparentsubstancefrom which acetylcholine is formed by enzymatic action ithe 2 vantageous source-.of -ascorbic"acid, (and enables this im-' portant vitamintonbe more widely used than heretofore possible. Moreover,-the presenceof thecholine factor in-ithe compound providesa single: source of two important vitamins; and the-compound is more desirable-from: thexstandpoi'nt :oftstabili-ty: and compatibility. with-othertherapeutic agents. thamthebasic constituents from which: it iS-fQrfi'lfidz 1 v Aocor dingMommas-invention.- choline ascorbate. iszprepared {byi'reaetings ascorbic-.zacid with a rchol'in'e basegzior I with reactants which form a choline base, in the presencelatter being of fundamental -importancefor-the-normalfunctioning of the nervous system; cholineis also con! sidered important in the maintenance of normalfat' trans-n port in the body and its therapeutic administrationi hasi proved invaluable in the prophylaxis'and: treatmehtlfof arteriosclerosis;

..Accor.dingly,..an object ofthepresentinvention; isto produce and to provide methods -of 'producin'g"a deriva tive of ascorbic"acid'which'is relatively.stablefunder; storage, conditions and is compatible with other constituents of therapeutic or nutrient compositions in which an ascorbic acid'factor is desirable, V

Another-"-object"of' the invention is to produce and providemethods' for producing a novel stablecompound containing'both ascorbic. acid and choline factors.

' Another *object ofthe invention is. to produce and provide" methods of producing" choline ascorbate.

Another obj'ectof' theinvention is to provide a stable solution containing an Lascorbioacid factor which is suitablefor parenteral use.

Another .objectof theinventionis. to provide a stable therapeuticor nutrient composition containing choline ascorbate. and vitamin. B-12 andfior other therapeutic agents. and vitamins It has been discovered that choline ascorbateypre pared. inaccordance with; the present invention, has a much better stability 1 in solution and greater compatibility with other therapeutic agents .and vitamins than ascorbic acid frompresently available sources. Accordingly,-the novel compound represents a new and ad-' ofwa suitabler'non reactive solventzmed-iumir The :solvent: med-iumimayirbeoanwof a number-erof vvell'knownf-com v mercial soliventsz-includingwvater;thesl ower alcohols; "and: non-polar solvents such as acetone, lactones, and esters. While the choice of solvent medium is not particularly critical, meth an'ohis preferred'because"of its'ready availability and ease of separation by distillation at low temperatures.

A suitable choline base may .b e*prepared by the re action of trimethylamine, ethylenooxide, and water in known.- marines-and; athis :may be employed in the reaction withtascorbie acid-r Alternatively, thecholine base forming..-constituents-..-maybe reacted? directly with ascorbici acid, or In this; methodwthe. ascorbic acid. iSwfil'St neutrali-zedewith trimethylamine-and then further reacted with- -ethylene.-oxide.-.f- Therea'ctants: arepr'eferably employed irrrsubstantiallyi equimolar quantities and both reactionsnresulein..su-bstanflally quantitative yields of the mono=choline salt-of=ascorbic acid;

The use of a greater than equimolar quantity of choline or choline-forming*c'onstituents appears to have; no effect-upon the reaction other thanto decrease-the:- product -yield.

.The product is'a heavy viscous liquid having a molecularweightof 1 279.28. It is quite soluble in water, sorbitol; and" in other solvents and forms stable solutions eithefalone or inix'ed'with other therapeutic agents or vitamins :suchi as ivitamin "B -12, thiamine,Ihydrochlotide," riboflavin,..-- sodium pantoth enate and 1 niacinami'de i as hereinafter; dis closed.

Stable aqueous solutions suitable for therapeutic use may be prepared containing, for .example, sufficient choline ascorbate "to provide" from about "lOI'Smillia gramspercubid'centiineter to 5'00milligrarns or .more'of ascorbic acid (based upon an ascorbic acid'con't'ent of 5658% To these" solutionsmay be" added' .othe'r lvitamins in substantially any"'desir'e'd quantity, depending upon the extentsof -rtheir solubility inuthes solvent employed, without adverse effect by reason of the choline 1 ascorbatea and.without-materially decreasing the ashelf conjunction with the following specific examples. It:

should be understood however that these-examplesare given merely by way o thillustration, ia ndinot by way oflimitation.

EXAMPLE I 121.18 mgs. (1.0 mole) of-freshlypreparedcholine base is dissolved in 500 cc. of methanol. To. this is added 183f-gms. (1.04 mole)"of ascorbic acid which has been suspended'in ZOO 'cc; of' methanol. The solution is; treatedwith carbon"and filtered;v then vacuum distilled to: remove methanol and water; A. 'yieldv of '270,gms;"of. choline ascorbate (97.0%) was obtained.

3 EXAMPLE n 183 gms. (10.4 mole) of ascorbic acid is suspended in 300 cc. of methanol and neutralized with a solution of 60.0 gms. (1.0 mole) of the methylamine in 200 cc. of methanol, while using external cooling to maintain the temperature below 35 C. The reaction mixture is then transferred to a flask equipped with a brine-cooled condenser, and a solution of 48.4 gms. (1.10 mole) of ethylene oxide in 200 cc. of methanol is slowly added while maintaining the temperature between 4560 C. After addition is completed, the solution is carbon treated and filtered, then vacuum distilled to remove methanol. A yield of 270 gms. of choline ascorbate (97.0%) was obtained.

The product as produced by the above examples was determined by analyses to be a new compound, monocholine ascorbate, having the following formula:

The choline content was determined by precipitation as a Reineckate in an acid medium, dissolving the Reineckate salt in a solvent and determining the quantitative content spectrophotometrically at a wave length of 525 millimicrons. Ascorbic acid was determined by iodine titration. Nitrogen determinations were conducted by the classic Kjeldahl method. Moisture determinations were made by the Karl-Fischer technique. The results of one such analyses are listed in Table I below:

Table I Found Throretical As is Dry Basis Basis Choline Content -percent" 43. 39 38.0 43. 7 Ascorbic Acid Content. do 63.06 56. 8 65. 4 Total Nitrogen Content do 4. 35 5. 01 Moisture Content do 13. 1 pH Value (2.5 Solution) 6.0

It will be noted that the results of analyses compare favorably with the theoretical content, within the margin of experimental error.

The chemical and physical properties of the new compound are further summarized below:

Choline ascorbateC H O-;N

10. Reaction:

(pH 2.5% solution) 6.00.

The following examples are illustrative of a number of compositions that may be prepared from choline ascorbate, and further show theadvantageous features of stability and compatibility of this compound as compared with ascorbic acid.

4 EXAMPLE HI Table II STABILITY OF CHOLINE ASCORBAIE AT ACCELERATED STORAGE TEMPERATURE Solution N o 1 2 3 4 Ascorbic Acid Content mg./cc 1O 25 50 Choline Ascorbate used Ooased on 56.8% as Ascorbic Acid) grams- 1. 768 4. 404 8.808 17. 620 Solvent, Volume (0 Initial Ascorbic Acid Content Inga/cc 10. 0 25. 1 50. l 106. 6 Storage Assay-Ascorbic Acid Content mg./cc 9. 4 23. 9 49.0 105. 3 Percent Ascorbic Acid Loss on Storage 6. 0 4. 8 2. 2 1. 4

1 Water, q. s., 100 cc.

The above results indicate that choline ascorbate is stable in solution for 550 hours at 45 C., these conditions being comparable to one year shelf life at normal temperatures.

EXAMPLE IV Duplicate solutions, comparable to 1, 2, 3 and 4 were prepared and 10 micrograms per cc. of vitamin 8-12 were added to ascertain whether these compounds were compatible. Initial assays were conducted and the samples were placed under accelerated storage conditions (temp. 45 C.) for 550 hours and then reassayed. The results of this experiment are shown in Table III:

Table III Solution N o 1 2 3 4 Ascorbic Acid Content as Choline Ascorbate mg./cc 10 25 50 100 B 12 added mcg./cc 10 10 10 10 Initial Ascor 0 Acid Content (by Iodine Titration) Ing./cc 10. 0 25. 9 60. 7 99. 3 Ascorbic Acid Content after storage at 45. C. for 550 hours -..rng./cc 9. 5 24. 3 49. 3 97.8 Percent Loss of Ascorbic Acid on Storage 5 6. 2 8. 8 15 Initial Vitamin B-12 Content by microbiological, assay) mcg./cc 10.9 10.9 10.9 10.9 Vitamin B-12 Content after Storage mcg./cc l1. 0 10. 4 9. 2 10. 6 Percent B-12 Loss on Storage None 4. 5 15. 5 2. 8

The results given in Table III indicate that choline ascorbate is compatible with vitamin B-12 and that thestability of the compounds is maintained within satisfactory'limits when the composition is stored at 45 C. for 550 hours, these conditions being equivalent to shelf life of one year at normal temperature.

EXAMPLE V A third test series was conducted by preparing solutions of choline ascorbate, equivalent in ascorbic acid content to 50 mg./cc. Solutions of commercially available ascorbic acid were also prepared in the same concentration. To these solutions were added 10 mcg. of

vitamin B-12, as well as other vitamins in the B-complex, namely, thiamine hydrochloride, riboflavin, sodium 1 Pius other factors of the B-complex. 9 70% Sorbltol. Water.

Table IV clearly illustrates that choline ascorbate offers a stable form of ascorbic acid and is compatible with vitamin B-12 and other factors of the B-complex. By comparison commercially available ascorbic acid is substantially completely incompatible with vitamin B-12 and the solutions are unstable and consequently impractical for therapeutic formulation.

Choline ascorbate in addition to its usage in applications wherein ascorbic acid or choline factors have been indicated, is believed to be of particular importance as a therapeutic agent in the management of arteriosclerosis. Thus, the action of the normal fat transport of choline to keep the fat mobilized and counteract stress on the arterial wall, in combination with the action of ascorbic acid on the ground substance of the arterial intima, provides an extremely efiective agent in the management of this impairment.

It will be apparent that applicant has provided a novel and useful new compound and compositions thereof, and has further provided methods for producing such novel compound. It will further be understood that various changes may be made in proportions of ingredients of the compositions disclosed, in the solvent media employed, and in other conditions and factors, without departing from the principles of the invention, or the scope thereof as set forth in the accompanying claims.

I claim:

1. A method for preparing choline ascorbate comprising reacting ascorbic acid in the presence of an inert solvent medium with a substantially equimolar quantity of a member selected from the group consisting of (I) choline and (II) the choline-forming reagents trimethylamine and ethylene oxide, and separating the solvent from the reaction product.

2. The method as defined in claim 1 wherein the solvent medium is methanol.

3. A method of preparing choline ascorbate comprising reacting ascorbic acid and choline in substantialy equimolar quantities in the presence of a solvent medium and separating the solvent from the reaction product by distillation.

4. A method for the preparation of choline ascorbate comprising neutralizing ascorbic acid with trimethylamine in the presence of a solvent, introducing ethylene oxide into the resulting reaction medium, and separating the solvent medium from the reaction product, the ascorbic acid, trimethylamine and ethylene oxide being employed in substantially equimolar proportions.

5. The method as defined in claim 4 in which the solvent medium is methanol, and the methanol is separated from the reaction product by distillation.

6. A method for the preparation of choline ascorbate comprising dissolving choline in methanol, adding a suspension of ascorbic acid in methanol to said solution the amount of ascorbic acid being substantially equimolar with respect to the choline, treating the resulting reaction mass with carbon, filtering the treated solution, and distilling methanol from said solution to recover choline ascorbate.

7. A method for the preparation of choline ascorbate comprising suspending ascorbic acid in methanol, neutralizing said suspension by the addition of a methanol solution of trimethylamine, adding a methanol solution of ethylene oxide to the resulting reaction mass the ascorbic acid, trimethylamine and ethylene oxide being employed in substantially equimolar proportions, treating the resultant reaction mass with carbon, filtering the treated solution from said carbon, and distilling said solution to remove methanol and recover choline ascorbate.

8. A viscous liquid having a specific gravity of approximately 1.32 at 25 C., and ascorbic acid content of approximately 43.7% as determined by iodine titration, a choline content of approximately 65.4% as determined by Reineckate precipitation, and a nitrogen content of approximately 5.01 by Kjeldahl determination, said choline and ascorbic acid components being combined as choline ascorbate.

9. A stable aqueous solution containing choline and ascorbic acid components in substantially equimolar proportions, said choline and ascorbic acid components being combined as choline ascorbate.

10. A vitamin composition containing choline and ascorbic acid components in substantially equimolar proportions, said choline and ascorbic acid components being combined as choline ascorbate.

11. A vitamin composition containing a B-complex vitamin, and choline and ascorbic acid components in substantially equimolar proportions, said choline and ascorbic acid components being combined as choline ascorbate.

12. A vitamin composition containing vitamin B-12, and choline and ascorbic acid components in substantially equimolar proportions, said choline and ascorbic acid components being combined as choline ascorbate.

References Cited in the file of this patent Bunatyan: Chem. Absts., vol. 37 (1943), pp. 6714 and 6715.

U. S. Dispensatory, 24th ed. (1947), p. 1400.

Howard: Modern Drug Encyclopedia, 5th ed. (1952), p. 1212. 

1. A METHOD FOR PREPARING CHOLINE ASCORBATE COMPRISING REACTING ASCORBIC ACID IN THE PRESENCE OF AN INERT SOLVENT MEDIUM WITH A SUBSTANTIALLY EQUIMOLAR QUANTITY OF A MEMBER SELECTED FROM THE GROUP CONSISTING OF (I) CHOLINE AND (II) THE CHOLINE-FORMING REAGENTS TRIMETHYLAMINE AND ETHYLENE OXIDE, AND SEPARATING THE SOLVENT FROM THE REACTION PRODUCT.
 6. A METHOD FOR THE PREPARATION OF CHOLINE ASCORBATE COMPRISING DISSOLVING CHOLINE IN METHANOL, ADDING A SUSPENSION OF ASCORBIC ACID IN METHANOL TO SAID SOLUTION THE AMOUNT OF ASCORBIC ACID BEING SUBSTANTIALLY EQUIMOLAR WITH RESPECT TO THE CHOLINE, TREATING THE RESULTING REACTION MASS WITH CARBON, FILTERING THE TREATED SOLUTION, AND DISTILLING METHANOL FROM SAID SOLUTION TO RECOVER CHOLINE ASCORBATE. 